Journal: Cell host & microbe

Article Title: A genomic toolkit for the mechanistic dissection of intractable human gut bacteria

doi: 10.1016/j.chom.2020.04.006

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Although our samples were not fully randomized across these variables, applying this correction substantially increased the correlation between metabolomic and phylogenetic distance and the profile similarity between the type strain isolates DSM 2243D, UCSF 2243, and ATCC 25559, which supported the value of the correction.

Techniques: Recombinant, Sequencing, Software

Journal: PLoS ONE

Article Title: A Robust High Throughput Platform to Generate Functional Recombinant Monoclonal Antibodies Using Rabbit B Cells from Peripheral Blood

doi: 10.1371/journal.pone.0086184

Figure Lengend Snippet: ( A ) Percentage of IgG-producing B-cell clones per total wells and ( B ) average IgG concentration over all IgG-positive wells after +/− pre-incubation in medium and +/− centrifugation post sorting. For each parameter 368 wells were analyzed. ( C ) Percentage of IgG-producing B-cell clones (ASCs) per total wells and ( D ) average IgG concentration over all IgG-positive wells after using +/− SAC in a dilution of 1∶20000 during B-cell cultivation. For each parameter 252 wells were analyzed. ( E ) Percentage of IgG-producing B-cell clones per total wells, ( F ) average IgG concentration over all IgG-positive wells, as well as percentage of antigen specific B-cell clones ( G ) per total wells and ( H ) per IgG-producing B-cell clones after +/− protein panning. For each parameter around 3500 wells were analyzed. The error bars represent the standard deviation. The cut off value of IgG-positive wells was >0.013 µg/ml IgG and of human IL1RL1-positive wells was>OD 0.195.

Article Snippet: 384-well MaxiSorp microtiter plates (Sigma-Aldrich, Nunc) were coated with 1 µg/ml goat anti-human IgG Fc fragment (Jackson Immuno Research) for at least 2 h. Thereafter the wells were blocked with PBS supplemented with 0.1% Tween®-20 and 2% BSA for 1 h. Recombinant human IL1RL1 Fc fusion protein (60 ng/ml, R&D Systems) was captured on a plate for 1 h. Dilutions of supernatants from rabbit ASCs or purified antibodies in PBS with 0.5% BSA and 0.05% Tween®-20 were incubated with the receptor protein for 1 h. Biotinylated human IL33 (PeproTech) was added for an additional hour to build up the complex.

Techniques: Clone Assay, Concentration Assay, Incubation, Centrifugation, Standard Deviation

Journal: PLoS ONE

Article Title: A Robust High Throughput Platform to Generate Functional Recombinant Monoclonal Antibodies Using Rabbit B Cells from Peripheral Blood

doi: 10.1371/journal.pone.0086184

Figure Lengend Snippet: Scatter Plots depicting the yield of the primary screening using all 7644 supernatants: ( A ) Human IL1RL1 binding (unit: optical density (OD)) versus IgG concentration; ( B ) human IL1RL1 binding versus cynomolgus IL1RL1 binding or versus ( C ) murine IL1RL1 binding. Scatter Plot showing the correlation of the biochemical inhibition assay with the cellular inhibition assay: ( D ) Threshold ≥40% inhibition, RSq: 0.36, ( E ) magnification of using the threshold of >90% inhibition, RSq: 0.9. The statistically confirmed cut off values for the calculation of the percentages were as follows: rabbit IgG-positive >0.013 µg/ml, human IL1RL1-positive >OD 0.195, human Fc-positive ≤OD 0.125, cynomolgus IL1RL1-positive >OD 0.184, murine IL1RL1-positive >OD 0.164. Green are the supernatants deriving from the pre-incubation only scenario and red are the SN after the protein panning step. The diamond, the circle and the cross indicate the three different animals. ( F ) Result of the two dimensional binding matrix identifying different binding epitopes on human IL1RL1. The colored numbers indicate different antigen specific antibodies. The black numbers describe the three antibody groups. The degree of antibody competition in the matrix is depicted by a 3-colour scale with green, black, red color indicating highest competition, mid or lowest competition, respectively.

Article Snippet: 384-well MaxiSorp microtiter plates (Sigma-Aldrich, Nunc) were coated with 1 µg/ml goat anti-human IgG Fc fragment (Jackson Immuno Research) for at least 2 h. Thereafter the wells were blocked with PBS supplemented with 0.1% Tween®-20 and 2% BSA for 1 h. Recombinant human IL1RL1 Fc fusion protein (60 ng/ml, R&D Systems) was captured on a plate for 1 h. Dilutions of supernatants from rabbit ASCs or purified antibodies in PBS with 0.5% BSA and 0.05% Tween®-20 were incubated with the receptor protein for 1 h. Biotinylated human IL33 (PeproTech) was added for an additional hour to build up the complex.

Techniques: Binding Assay, Concentration Assay, Inhibition, Incubation

Journal: International Journal of Molecular Sciences

Article Title: β-Casein A1 and A2 Genetic Variants and β-Casomorphin-7 in Raw Milk and Processed Milk Products

doi: 10.3390/ijms26178612

Figure Lengend Snippet: Proposed mechanism of β-casomorphin-7 (BCM-7) release from β-casein A1 variant during gastrointestinal digestion. Variant A2 does not generate BCM-7.

Article Snippet: After a brief homogenization, raw milk or dairy products were tested for the presence of A1 and A2 beta-casein variants using the Bovine A1 (or A2) Beta-Casein ELISA Kit (Biosensis Pty Ltd., Thebarton, Australia).

Techniques: Variant Assay

Journal: International Journal of Molecular Sciences

Article Title: β-Casein A1 and A2 Genetic Variants and β-Casomorphin-7 in Raw Milk and Processed Milk Products

doi: 10.3390/ijms26178612

Figure Lengend Snippet: Workflow of experiments leading to measure the content of β-casein variants A1 and A2 as well as BCM-7 in the raw milk and popular processed dairy products. By A–G letters, 7 dairy plants are abbreviated.

Article Snippet: After a brief homogenization, raw milk or dairy products were tested for the presence of A1 and A2 beta-casein variants using the Bovine A1 (or A2) Beta-Casein ELISA Kit (Biosensis Pty Ltd., Thebarton, Australia).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: β-Casein A1 and A2 Genetic Variants and β-Casomorphin-7 in Raw Milk and Processed Milk Products

doi: 10.3390/ijms26178612

Figure Lengend Snippet: Relative content of β-casein A1 and A2 variants in raw milk and processed dairy products (pasteurized milk, UHT milk, and milk powder). Bars represent mean ± SD. Different superscript letters indicate significant differences ( p < 0.01) among product types. Note: in UHT milk, difference between the content of A1 and A2 variants did not reach statistical significance ( p > 0.05) due to high variability among samples. Different capital letters A and B indicate statistical significance between means at level p < 0.01.

Article Snippet: After a brief homogenization, raw milk or dairy products were tested for the presence of A1 and A2 beta-casein variants using the Bovine A1 (or A2) Beta-Casein ELISA Kit (Biosensis Pty Ltd., Thebarton, Australia).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: β-Casein A1 and A2 Genetic Variants and β-Casomorphin-7 in Raw Milk and Processed Milk Products

doi: 10.3390/ijms26178612

Figure Lengend Snippet: Mean values and standard deviation (± SD) of β-casein variant A1 (left) and variant A2 (right) contents in raw milk obtained from cows with identified CSN2 genotype. Statistically significant differences ( p < 0.01) are indicated with different superscript letters. Different capital letters A and B indicate statistical significance between means at level p < 0.01.

Article Snippet: After a brief homogenization, raw milk or dairy products were tested for the presence of A1 and A2 beta-casein variants using the Bovine A1 (or A2) Beta-Casein ELISA Kit (Biosensis Pty Ltd., Thebarton, Australia).

Techniques: Standard Deviation, Variant Assay